Using Dihydropyridine-Release Strategies to Enhance Load Effects in Engineered Human Bone Constructs

Abstract

We report on the development of a novel biodegradable scaffold capable of enhancing mechanical signals for tissue-engineering applications. It has been shown that mechanotransduction enhances bone formation in vitro and in vivo; in tissue-engineering applications, this phenomenon is exploited through the use of mechanical bioreactors to generate bone tissue. The dihydropyridine agonist Bay K8644 (Bay) acts to increase the opening time of mechanosensitive voltage-operated calcium channels (VOCCs), specifi- cally L-type VOCCs, which are known to play a fundamental role in the early mediation of mechanotransduction. We have produced porous 3-dimensional, Bay-encapsulated biodegradable poly(L-lactide) acid scaffolds using a solvent-casting and salt-leaching technique. The effects of the released Bay on osteoid production and mineralization in human bone cell-seeded constructs following incubation in a perfusion-compression bioreactor in vitro was investigated using Western blotting techniques and a calcium assay protocol developed in our lab. Our newly developed scaffolds act by slowly releasing the calcium channel agonist Bay K8644 as observed using ultraviolet spectroscopy, maintaining the open state of mechanosensitive VOCCs responding to load, which augments the load signal at sites of strain across the scaffold. Our results demonstrate that, in the presence of physiological loading regimes in vitro, release of Bay enhances collagen I protein production and osteoid calcification more than non-Bay control constructs do. Osteopontin and alpha2delta1 VOCC subunit protein levels were also higher as a result of perfusion-compression conditioning. These results indicate that Bay-encapsulated scaffolds can be used in the presence of load to enhance the production of load-bearing engineered tissue.

Biomechanical evaluation of porous biodegradable scaffolds for revision knee arthroplasty

“…This study was focused on the mechanical failure of the scaffold, but the strain distribution gives some insight into the osteogenic response that can be expected. It has indeed been reported that scaffold strain of the order of 1% promotes cell ingrowth and extracellular matrix synthesis (Wood et al 2006).…”

“…These conclusions will have to be confirmed by a more realistic model of the tibia and implant, which should also provide a better estimation of the osteogenic level of the strain. Whatever the mechanisms at the cellular level, strain level within the scaffold matrix is indeed crucial in the process of bone ingrowth into such biodegradable polymer scaffolds (Wood et al 2006;Duty et al 2007). …”

Biomechanics in bone tissue engineering

“…New designs, such as the biaxial rotating bioreactor (Zhang et al 2009), are continuously developed and are mostly based on empirical approaches, with the exception of those designed based on theoretical developments (Singh et al 2005) or the description of local strains sensed by the cells (Pfeiler et al 2008). To increase the responsiveness of the mechanically stimulated cells in a bioreactor, mechanical stimulation was combined with drug supplementation, which has an effect in increasing the opening time of mechanosensitive voltage-operated calcium channel (Wood et al 2006). A positive effect using this strategy was observed for the calcification of the osteoid.…”

Correlating cell morphology and osteoid mineralization relative to strain profile for bone tissue engineering applications

“…For example, the dihydropyridine agonist Bay K8644 has been encapsulated and released from mechanically robust scaffolds, acting to augment the opening state of L-type voltageoperated calcium channels (VOCC) following their activation via a controlled loading regime ( Yang et al 2002). The effects of Bay K8644-encapsulated scaffolds on bone tissue formation in the presence of load include increased collagen I production and osteoid mineralization within constructs ( Wood et al 2006b). Thus, mechanical conditioning of cell-seeded constructs, often in the presence of biochemicals, is attracting a great deal of attention due to reports of accelerated tissue production in three-dimensional scaffolds and matrices.…”